The invention relates to the identification of expression profiles and the nucleic acids involved in breast cancer, and to the use of such expression profiles and nucleic acids in diagnosis and prognosis of breast cancer. The invention further relates to methods for identifying and using candidate agents and/or targets which modulate breast cancer.
Breast cancer is a significant cancer in Western populations. It develops as the result of a pathologic transformation of normal breast epithelium to an invasive cancer. There have been a number of recently characterized genetic alterations that have been implicated in breast cancer. However, there is a need to identify all of the genetic alterations involved in the development of breast cancer.
Imaging of breast cancer for diagnosis has been problematic and limited. In addition, dissemination of tumor cells (metastases) to locoregional lymph nodes is an important prognostic factor; five year survival rates drop from 80 percent in patients with no lymph node metastases to 45 to 50 percent in those patients who do have lymph node metastases. A recent report showed that micrometastases can be detected from lymph nodes using reverse transcriptase-PCR methods based on the presence of mRNA for carcinoembryonic antigen, which has previously been shown to be present in the vast majority of breast cancers but not in normal tissues. Liefers et al., New England J. of Med. 339(4):223 (1998).
While academia and industry has made an effort to identify novel sequences, there has not been an equal effort exerted to identify the involvement of such sequences in disease states. For example, databases show the sequence for accession number AF183810, which encodes a polypeptide corresponding to a transcription factor, TRPS1. TRPS1 has been shown to contain multiple zinc-finger motifs and has been implicated in tricho-rhino-phalangeal syndrome type I, a malformation syndrome with craniofacial and skeletal abnormalities (Morneni, P. et al., Nat. Genet, 24:71-74 (2000)). However, no role in cancer has been ascribed to this protein until the present disclosure.
Thus, methods that can be used for diagnosis and prognosis of breast cancer would be desirable. Accordingly, provided herein are methods that can be used in diagnosis and prognosis of breast cancer. Further provided are methods that can be used to screen candidate bioactive agents for the ability to modulate breast cancer. Additionally, provided herein are molecular targets for therapeutic intervention in breast and other cancers.
The present invention provides methods for screening for compositions which modulate breast cancer.
In one aspect, a method of screening drug candidates comprises providing a cell that expresses an expression profile gene or fragments thereof. Preferred embodiments of the expression profile gene as described herein include the sequence comprising BFA4 or a fragment thereof. The method further includes adding a drug candidate to the cell and determining the effect of the drug candidate on the expression of the expression profile gene.
In one embodiment, the method of screening drug candidates includes comparing the level of expression in the absence of the drug candidate to the level of expression in the presence of the drug candidate, wherein the concentration of the drug candidate can vary when present, and wherein the comparison can occur after addition or removal of the drug candidate. In a preferred embodiment, the cell expresses at least two expression profile genes. The profile genes may show an increase or decrease.
Also provided herein is a method of screening for a bioactive agent capable of binding to a breast cancer modulating protein (BCMP) or a fragment thereof, the method comprising combining the BCMP or fragment thereof and a candidate bioactive agent, and determining the binding of the candidate agent to the BCMP or fragment thereof in a preferred embodiment, the BCMP is BFA4.
Further provided herein is a method for screening for a bioactive agent capable of modulating the bioactivity of a BCMP or a fragment thereof. In one embodiment, the method comprises combining the BCMP or fragment thereof and a candidate bioactive agent, and determining the effect of the candidate agent on the bioactivity of the BCMP or the fragment thereof. In a preferred embodiment the BCMP is BFA4.
Also provided herein is a method of evaluating the effect of a candidate breast cancer drug comprising administering the drug to a transgenic animal expressing or over-expressing a BCMP or a fragment thereof, or an animal lacking a BCMP for example as a result of a gene knockout in a preferred embodiment, the BCMP is BFA4.
Additionally, provided herein is a method of evaluating the effect of a candidate breast cancer drug comprising administering the drug to a patient and removing a cell sample from the patient The expression profile of the cell is then determined. This method may further comprise comparing the expression profile to an expression profile of a healthy individual.
Furthermore, a method of diagnosing breast cancer is provided. The method comprises determining the expression of a gene which encodes BFA4 or a fragment thereof in a first tissue type of a first individual, and comparing this to the expression of the gene from a second unaffected individual. A difference in the expression indicates that the first individual has breast cancer.
In another aspect, the present invention provides an antibody which specifically binds to BFA4, or a fragment thereof. Preferably the antibody is a monoclonal antibody. The antibody can be a fragment of an antibody such as a single stranded antibody as further described herein, or can be conjugated to another molecule. In one embodiment, the antibody is a humanized antibody.
In one embodiment a method for screening for a bioactive agent capable of interfering with the binding of BFA4 or a fragment thereof and an antibody which binds to said BFA4 or fragment thereof is provided. In a preferred embodiment, the method comprises combining BFA4 or a fragment thereof, a candidate bioactive agent and an antibody which binds to said BFA4 or fragment thereof. The method further includes determining the binding of said BFA4 or fragment thereof and said antibody Wherein there is a change in binding, an agent is identified as an interfering agent The interfering agent can be an agonist or an antagonist. Preferably, the antibody as well, as the agent inhibits breast cancer.
In one aspect of the invention, a method for inhibiting the activity of a breast cancer modulating protein are provided. The method comprises binding an inhibitor to the protein. In a preferred embodiment, the protein is BFA4.
In another aspect, the invention provides a method for neutralizing the effect of a breast cancer modulating protein. The method comprises contacting an agent specific for the protein with the protein in an amount sufficient to effect neutralization. In a preferred embodiment, the protein is BFA4.
In a further aspect, a method for treating or inhibiting breast cancer is provided. In one embodiment, the method comprises administering to a call a composition comprising an antibody to BFA4 or a fragment thereof. In one embodiment, the antibody is conjugated to a therapeutic moiety. Such therapeutic moieties include a cytotoxic agent and a radioisotope. The method can be performed in vitro or in vivo, preferably in vivo to an individual. In a preferred embodiment the method of inhibiting breast cancer is provided to an individual with such cancer.
As described herein, methods of treating or inhibiting breast cancer can be performed by administering an inhibitor of BFA4 activity to a cell or individual. In one embodiment a BFA4 inhibitor is an antisense molecule to a nucleic acid encoding BFA4.
Moreover, provided herein is a biochip comprising a nucleic acid segment which encodes BFA4, or a fragment thereof, wherein the biochip comprises fewer than 1000 nucleic acid probes. Preferably at least two nucleic acid segments are included.
Also provided herein are methods of eliciting an immune response in an individual. In one embodiment a method provided herein comprises administering to an individual a composition comprising BFA4 or a fragment thereof. In another aspect, said composition comprises a nucleic acid comprising a sequence encoding BFA4 or a fragment thereof.
Further provided herein are compositions capable of eliciting an immune response in an individual. In one embodiment a composition provided herein comprises BFA4 or a fragment thereof and a pharmaceutically acceptable carrier. In another embodiment, said composition comprises a nucleic acid comprising a sequence encoding BFA4 or a fragment thereof and a pharmaceutically acceptable carrier.
Other aspects of the invention will become apparent to the skilled artisan by the following description of the invention.